ABSTRACT
The development of oligomeric glucagon-like peptide-1 (GLP-1) and GLP-1-containing coagonists holds promise for enhancing the therapeutic potential of the GLP-1-based drugs for treating type 2 diabetes mellitus (T2DM). Here, we report a facile, efficient, and customizable strategy based on genetically encoded SpyCatcher-SpyTag chemistry and an inducible, cleavable self-aggregating tag (icSAT) scheme. icSAT-tagged SpyTag-fused GLP-1 and the dimeric or trimeric SpyCatcher scaffold were designed for dimeric or trimeric GLP-1, while icSAT-tagged SpyCatcher-fused GLP-1 and the icSAT-tagged SpyTag-fused GIP were designed for dual GLP-1/GIP (glucose-dependent insulinotropic polypeptide) receptor agonist. These SpyCatcher- and SpyTag-fused protein pairs were spontaneously ligated directly from the cell lysates. The subsequent icSAT scheme, coupled with a two-step standard column purification, resulted in target proteins with authentic N-termini, with yields ranging from 35 to 65 mg/L and purities exceeding 99%. In vitro assays revealed 3.0- to 4.1-fold increased activities for dimeric and trimeric GLP-1 compared to mono-GLP-1. The dual GLP-1/GIP receptor agonist exhibited balanced activity toward the GLP-1 receptor or the GIP receptor. All the proteins exhibited 1.8- to 3.0-fold prolonged half-lives in human serum compared to mono-GLP-1 or GIP. This study provides a generally applicable click biochemistry strategy for developing oligomeric or dual peptide/protein-based drug candidates.
Subject(s)
Click Chemistry , Glucagon-Like Peptide 1 , Glucagon-Like Peptide 1/chemistry , Humans , Receptors, Gastrointestinal Hormone/agonists , Receptors, Gastrointestinal Hormone/chemistry , Receptors, Gastrointestinal Hormone/metabolism , Drug Design , Diabetes Mellitus, Type 2/drug therapy , Gastric Inhibitory Polypeptide/chemistry , Gastric Inhibitory Polypeptide/pharmacology , Glucagon-Like Peptide-1 Receptor/agonistsABSTRACT
The prevalence of leprosy fell below 1 per 100,000 in Shandong province in 1994. Since then, a few incident leprosy cases have been reported each year over the past 10 years. In order to explore whether or not the reduction in case detection in the province was due to the stopping of active case-finding activities, a rapid village survey was conducted in a formerly endemic county, using skin disease service. Ninety-one villages in 23 townships, covering a total population of 104,885, were surveyed. No leprosy case was detected. The results of the survey, along with other evidences, such as higher MB/PB ratio among newly detected cases and late onset of the disease, suggest that leprosy is dying out as a disease in the county as well as in Shandong. The efforts of the leprosy control programme should shift to other services such as prevention of disability and care for the disabled.
